Abstract

Based on amino acid sequence information from purified mitochondrial thymidine kinase (TK2), a cDNA of 1930 bp was cloned, containing an open reading frame encoding 232 amino acid residues starting with the N-terminal sequence determined from the native human protein preparation, Northern blot analysis with the cDNA coding region demonstrated several TK2 mRNAs, with 2 and 4 kb forms present in many tissues. We also characterised N-terminally truncated (starting at position 18) human TK2 with pharmacologically important antiviral and cytostatic nucleoside analogues. Results were highly similar to those with the native TK2 preparation. The anti-leukaemic drug arabinosyl cytosine is phosphorylated. The antitumour drug difluorodeoxycytidine and its metabolite difluorodeoxyuridine are good substrates, with K-m values of 66 and 29 mu M, respectively, and a relative V-max of 0.6 compared to that of thymidine. Negative cooperativity was found with thymidine and the anti-HIV drug 3'-azidothymidine, but the reaction followed Michaelis-Menten kinetics with deoxycytidine, arabinosyl cytosine, and arabinosyl thymine. The results demonstrate a broad substrate specificity and complex kinetics, and suggest a role for TK2 in the activation of chemotherapeutic nucleoside analogues. (C) 1999 Federation of European Biochemical Societies.

Keywords

cloning; thymidine kinase; antiviral nucleoside; cytostatic nucleoside; metabolic conversion; mitochondrial enzyme

Published in

FEBS Letters
1999, Volume: 443, number: 2, pages: 170-174

SLU Authors

• Wang, Liya

  • Department of Veterinary Medical Chemistry, Swedish University of Agricultural Sciences
    

• Herrström Sjöberg, Anita

  • Department of Veterinary Medical Chemistry, Swedish University of Agricultural Sciences
    

• Eriksson, Staffan

  • Department of Veterinary Medical Chemistry, Swedish University of Agricultural Sciences
    

UKÄ Subject classification

Biochemistry and Molecular Biology


Publication identifier

DOI: https://doi.org/10.1016/S0014-5793(98)01711-6

Permanent link to this page (URI)

https://res.slu.se/id/publ/53118